. PROJECT SUMMARY (See instructions): The Flow Cytometry and Cellular imaging Facility (FCCIF) was established to provide access to state-of-the art cell analysis technology for MD Anderson investigators, and provides cell sorting, analytical flow cytometry, cellular imaging and custom monoclonal antibody (mAb) conjugations to its users. The Core provides researchers with technical expertise in instrument operation, assay development, data acquisition and various analysis techniques. Analytical flow cytometry is an indispensable tool for the study of all aspects of cell biology, including protein expression, cell proliferation and differentiation, cell signaling pathways, enzyme activity, gene regulation, ceil lineage, apoptosis, autophagy and chemotherapeutic resistance. The Core has recently acquired a DVS CyTOF Mass Cytometer, enabling the detection and characterization of up to 100 molecular markers at the single cell level. This instrument represents a transformational technology enabling the detection and characterization of rare and mixed cell populations on the single cell level. Cell sorting. Cell isolation for culture and further characterization is performed via droplet-based sorting, which isolates a wide variety of cells based on combinations of antibody-based stains, fluorescent protein expression, and viability indicators. Imaging. The Core offers researchers tools and techniques for image acquisition, SD-reconstruction, and time-series observation as well as a variety of image processing and analysis functions via laser scanning cytometry and confocal microscopy and also offers multispectral, epifluorescent, and colorimetric microscopy. Custom mAb conjugations. Antibody conjugation is a new service of the FCCIF that provides conjugates with fluors and tags that are not commercially available. The FCCIF uses 24 major instrument systems supporting the research of-345 investigators who hold 13 POIs, 112 ROIs, and 9 P50 SPOREs. Peer-reviewed investigators account for 94% of the utilization, and 35% of total cost is requested from the CCSG. Over the past 5 years, the FCCIF has performed more than 50,000 hours of service, representing a 125% increase over the prior grant period. Over the past 5 years, the FCCIF has facilitated publication of 408 reports, with 67% in journals with an impact factor >5 and 22% with an impact factor >10. In the future, the FCCIF will continue to develop the use of the current instrumentation, including the DVS Sciences CyTOF, and novel analysis programs, including the SPADE algorithm. Older equipment will be replaced, and an Amnis Image Stream, a Vectra 2 automated multispectral imaging system and single-cell analysis systems such as Fluidigm's BioMark may be added.